Mycobacterium tuberculosis: Macrophage Takeover and Modulation of Innate Effector Responses

Macrophages mediate the first line of defense in the host against various intracellular pathogens. They are armed with several immune-effector mechanisms to detect and combat pathogens. However, intracellular pathogens have developed strategies to overcome the macrophage protective immune responses and colonize inside the macrophages. Tuberculosis (TB), both pulmonary and extrapulmonary, is an infectious disease of global concern caused by Mycobacterium tuberculosis. M. tuberculosis is a highly successful pathogen and has acquired various strategies to downregulate critical innate-effector immune responses of macrophages such as phagosome-lysosome fusion, antigen presentation, autophagy, and inhibition of reactive oxygen (ROI) and reactive nitrogen (RNI) species to ensure its longer survival inside the macrophages. In addition to these, the bacilli also modulate T cell immune response which can help the bacilli to survive inside the host for a long time. In this chapter, we focus to describe important macrophage innate defense mechanisms and the signaling that can influence T cell adaptive response and the strategies adopted by the bacilli to exploit these signaling cascades to favor its replication and persistence inside the macrophages for establishing a productive infection

Mycobacteria modulate host epigenetic machinery by Rv1988 methylation of a non-tail arginine of histone H3

Mycobacteria are successful pathogens that modulate the host immune response through unclear mechanisms. Here we show that Rv1988, a secreted mycobacterial protein, is a functional methyltransferase that localizes to the host nucleus and interacts with chromatin. Rv1988 methylates histone H3 at H3R42 and represses the genes involved in the first line of defence against mycobacteria. H3R42me2, a non-tail histone modification, is present at the entry and exit point of DNA in the nucleosome and not within the regulatory sites in the N-terminal tail. Rv1988 deletion in Mycobacterium tuberculosis reduces bacterial survival in the host and experimental expression of M. tuberculosis Rv1988 in non-pathogenic Mycobacterium smegmatis negatively affects the health of infected mice. Thus, Rv1988 is an important mycobacterial virulence factor, which uses a non-canonical epigenetic mechanism to control host cell transcription

Learning epigenetic regulation from mycobacteria

In a eukaryotic cell, the transcriptional fate of a gene is determined by the profile of the epigenetic modifications it is associated with and the conformation it adopts within the chromatin. Therefore, the function that a cell performs is dictated by the sum total of the chromatin organization and the associated epigenetic modifications of each individual gene in the genome (epigenome). As the function of a cell during development and differentiation is determined by its microenvironment, any factor that can alter this microenvironment should be able to alter the epigenome of a cell. In the study published in Nature Communications (Yaseen [2015] Nature Communications 6:8922 doi: 10.1038/ncomms9922), we show that pathogenic Mycobacterium tuberculosis has evolved strategies to exploit this pliability of the host epigenome for its own survival. We describe the identification of a methyltransferase from M. tuberculosis that functions to modulate the host epigenome by methylating a novel, non-canonical arginine, H3R42 in histone H3. In another study, we showed that the mycobacterial protein Rv2966c methylates cytosines present in non-CpG context within host genomic DNA upon infection. Proteins with ability to directly methylate host histones H3 at a novel lysine residue (H3K14) has also been identified from Legionella pnemophilia (RomA). All these studies indicate the use of non-canonical epigenetic mechanisms by pathogenic bacteria to hijack the host transcriptional machinery

Optimization of ultrasonic-assisted extraction of eugenol-rich fraction from basil leaves: Characterization of extract for phenols, flavonoids and antioxidant activity

In this study, optimization of ultrasonic assisted extraction (UAE) of eugenol-rich fraction from O. basilicum was performed. Box-Behnken design of response surface methodology was applied with three independent variables viz. solvent concentration (25–75%), amplitude (70–90%) and sonication time (7–12 min). The effects of these three independent variables were studied on extract yield, total phenolic content, total flavonoid content, anti-oxidant activity and eugenol content present in O. basilicum. Significant (p≤0.05) effects of all the independent variables were recorded in all the product responses. Extract yield and total phenolic content ranged from 74.36–55.48% and 23.45–10.25 mgGAE/g, respectively while as total flavonoid content and anti-oxidant activity was 0.082–0.009 mg/g and 71.21–25.33% respectively and eugenol content of 0.846–0.207 mg/g was recorded for all the treatments. The optimum processing conditions obtained by numerical optimization for solvent concentration (A), amplitude (B) and sonication time (C) were 52.48%, 90% and 7.75 min, respectively. Eugenol content of 41.44% was further validated through GC-MS/MS in extract developed after following the optimal condition. Conclusively, RSM was found to be successful for optimization of UAE process of eugenol rich fraction from basil leaves. The results confirmed the success of using ethanol as a safe solvent for UAE of eugenol from basil leaves

Orange to red emissive aldehyde substituted donor-π-acceptor phenothiazine derivatives: Optoelectronic, DFT and thermal studies

A new class of probes was synthesized using a simple and efficient synthetic protocol. These compounds (PTZ-6(a-e)) have the phenothiazine (PTZ) moiety as the electron donor (D) and substituted aldehydes along with the acrylonitrile group, which acts as the electron acceptor (A), thus making D-π-A push-pull system. The structures of the newly synthesized series of small organic target molecules PTZ-6(a-e) were investigated and confirmed by spectros-copic techniques. The optical/solvatochromic properties were studied in detail by UV-vis absorption and fluorescence spectroscopy, because the molecules have shown good solubility in organic solvents. The density functional theory (DFT) model with the CAM-B3LYP function is utilized to study the photophysical properties of the probes, as these probes exhibited orange-to-red emission. Optical band gap values ranged from 2.32 to 2.50 eV, and these probes exhibited good thermal stability with a melting temperature of 136 to 198 °C and a T5d temperature range from 335 to 354 °C. The cyclic voltammetry study confirms that the Eoxonset values of the target compounds are 0.80 eV. The quantum yields (Φ) of the probes are measured experimentally in ethanol and the Stokes shifts are observed to be in the range of 4846-9430 cm-1. The results displayed that novel (D-A-D) chromophores could play an important role in organic optoelectronics

Proteasome-dependent truncation of the negative heterochromatin regulator Epe1 mediates antifungal resistance

Epe1 histone demethylase restricts H3K9-methylation-dependent heterochromatin, preventing it from spreading over, and silencing, gene-containing regions in fission yeast. External stress induces an adaptive response allowing heterochromatin island formation that confers resistance on surviving wild-type lineages. Here we investigate the mechanism by which Epe1 is regulated in response to stress. Exposure to caffeine or antifungals results in Epe1 ubiquitylation and proteasome-dependent removal of the N-terminal 150 residues from Epe1, generating truncated tEpe1 which accumulates in the cytoplasm. Constitutive tEpe1 expression increases H3K9 methylation over several chromosomal regions, reducing expression of underlying genes and enhancing resistance. Reciprocally, constitutive non-cleavable Epe1 expression decreases resistance. tEpe1-mediated resistance requires a functional JmjC demethylase domain. Moreover, caffeine-induced Epe1-to-tEpe1 cleavage is dependent on an intact cell-integrity MAP kinase stress signalling pathway, mutations in which alter resistance. Thus, environmental changes elicit a mechanism that curtails the function of this key epigenetic modifier, allowing heterochromatin to reprogram gene expression, thereby bestowing resistance to some cells within a population. H3K9me-heterochromatin components are conserved in human and crop plant fungal pathogens for which a limited number of antifungals exist. Our findings reveal how transient heterochromatin-dependent antifungal resistant epimutations develop and thus inform on how they might be countered